Process for preparing metal salts of penicillin



Patented Dec. 11, 1951 PROCESS FOR. PREPARING METAL SAL-'IS F PENICILLINMax Tishlei', Westfild, N. 1., assignor to Merck & Co., Inc., Rahway, N.J., a corporation of New Jersey No Drawing-".1 Original applicationSeptember 11, 1946, Serial N0. 696,350. Divided and this applicationFebruary 25, 1950, Serial No. 146,410

4 Claims. '(Cl. 260-= 239.1)

This application is a, division of my pending joint -application, Cramand Tishler, 'Serial No. 696,350, filed September 11, 1946, now UnitedStates Patent No. 2,542,796.

This invention relates to the isolation and. purification of penicillinG from a mixture of various types of penicillin. More particularly, itis concerned with the preparation of n'ovelder-ivatives of penicillinG'vvhich permit their ready separation.

It has been established with reasonable certainty that penicillin G,Sometimes referred to as benzyl penicillin, has the structural formulaBro [N as v (EH-(i=0 I IHC O-R wherein R represents the benzyl group('CH2-'C6H5) Other forms of penicillin "are of similar structurediffering only in that the benzyl group is replaced by othermon'ovalen't organic radicals, generally radicals having five or morecarbon atoms. Penicillins, such as penicillin F, wherein R is A-rp'en'tenyl (--CHz-CH=CH-CH2CH2) dihydropenicillinE, wherein Risn-amyl;penicillin X, wherein R is p-hydroxy benzyl and penicillin K, wherein Ris n-heptyl have been identified. Although both saturated andunsaturated hydrocarbon radicals at R in the above formula give productshaving penicillin activity, it has been found that penicillin G is moreeflicacious than some of the other forms in therapy.

Penicillin is produced commercially as a mixture of these various typescontaining pigments and extraneous material. Since penicillin G is oneof the most active forms, it is desirable that it be separated from theless active types as well as extraneous impurities.

In accordance with one method of isolating penicillin G, described in apenicillin report submitted by the Heyden Chemical Corporation to the O.S. R. D. (O. S. R. D. No. H-II, May 22, 1944, and O. S. R. D. No. H-III,June 15, 1944), an ether solution of triethylamine was added to an ethersolution of crude penicillin and a precipitation of impurities effected.Upon further addition of an ether solution of triethylamine to thepenicillin solution, a mixture of oil and crystals formed.

sodium salt was recorded. it consisted of dissolving the triethylam'in'esalt in water, acidifying, extracting into ether, and back into waterbythe 'portio'nwi'se addition of sodium hydroxide. This mixture was thenfrozen, dried under vacuum and recrystallized from butanol and water.The'above procedure; however,-does not give high yields of penicillin Gand therefore not economical for manufacture.

It is an object of this invention to provide a practical process for theisolation and purification'o'f penicillin G from a mixture, which can bereadily carried out and does not result in significant losses ofpenicillin. V

This improved method for isolation and purification of penicillin Gincludes the steps of treating an organic solvent solution of penicillinacid with an organic solvent solution of an N-subst'ituted heteroc'yclicamine to form the tertiary amine salt of penicillin G, recovering thelatter "compound'and converting the same to an alkali or alkaline earthmetal salt of essentially pure penicillin G. 'The firs'tstep of the"overall process, i. e., the formation of the tertiary amine salt isfullydisclosed and claimed in said pending application of Cram andTishlii, SerialNO. 696,350, filed September 1 1 1946, typical tertiaryamine salts prepared according to said process including the N-methylpiperidine, N-ethylpiperidine, N-methylmorpholine andN-ethylmorpholine salts of penicillin G.

Regarded in certain of its broader aspects, the process-in accordancewith the present invention comprises converting a tertiary amine salt ofpenicillin G to an alkali metal or alkaline earth metal salt ofpenicillin G by dissolving the tertiary amine salt in an aqueous organicsolvent mixture, reacting the latter mixture with an alkali formingmetal hydroxide, and recovering the corresponding alkaline salt ofpenicillin G from the aqueous phase. By the term alkali forminghydroxide as used here is meant any alkali metal or alkaline earth metalhydroxide.

By way of illustration, conversion of the N- ethylpiperidine salt ofpenicillin G to sodium penicillin G or calcium penicillin G isaccomplished by dissolving the N-ethylpiperidine salt in water andadding amyl acetate. Suificient dilute sodium hydroxide or calciumhydroxide is added to the above mixture to adjust the pH of the solutionto approximately 6.5. The alkali is added at such a rate that thetemperature of the reaction does not exceed 3 C. In this manner thesodium or ca cium penicillin G is extracted into the aqueous layer andthe N-ethylpiperidine is extracted into the amyl acetate. The two layersare separated. The aqueous layer is washed with amyl acetate. recoveredfrom the aqueous solution by freezing the solution and drying undervacuum. An essentially pure product is obtained. N-ethylpiperidine issoluble in amyl acetate, and can be recovered from the amyl acetatelayer.

Th amo-r"h011s sodium penici lin G t us tained can be crystallized bystirring the product ith butanol until solution is accom lished.Crystallization begins almost immediately.

- The following example illustrates a method of carrying out the presentinvention. It is to be understood that this example is given by way ofillustration and not of limitation.

Example The N-ethylpiperidine salt of penicillin G is converted to puresodium penicillin G by dissolving 6.0 g. of the pure N-ethylpiperidinesalt in 35.8 cc. of cold water (carbon dioxide free) and covering theaqueous layer with 6'? cc. of cold amyl acetate. The mixture is stirredat C. and treated with 13.5 cc. of 0.9955 N sodium hydroxide (carbonatefree). The alkali is added at such a rate that the temperature does notexceed 3 C. The two layers are separated and the aqueous layer extractedtwo times with 35 cc. of cold amyl acetate. The aqueous layer which hasa pH of 7, is frozen and dried under vacuum to give 4.78 g. ofessentially pure colorless sodium penicillin G. This represents 100% oftheory. The product had an optical rotation (a)25 =292.5 when 100 mg.was measured in 25 cc. water.

Bioassay showed S. Marcus 1652 u./mg.

If calcium hydroxide is used in place of sodium hydroxide, pure calciumpenicillin G is obtained.

The above represents a 76% overall recovery of activity from clinicalpenicillin as sodium penicillin G.

Anal. calcd. for C16H17N2SO4N8.

Found C, 54.08; H, 5.02; N, 7.74

Amorphous sodium penicillin is.

The N-ethylpiperidine is recovered from the amyl acetate layer by adding2.5 N hydrochloric acid to the amyl acetate until acid to moist Congo.The layers are separated and the aqueous layer made alkaline with 30%sodium hydroxide. The N-ethylpiperidine is separated, dried overpotassium hydroxide pellets and distilled at atmospheric pressure.

Crystallization of the sodium penicillin G obtained above isaccomplished by stirring a mixture of 5.0 g. of the product and 15 cc.of butanol until the sodium penicillin G dissolves and then crystallizesfrom solution. The mixture is cooled at 0 C. for five hours, filtered,slurried two times with a minimum volume of acetone and dried in vacuo.4.66 g. (93.2% recovery) of crystalline sodium penicillin G is obtained.

Various changes and modifications in the foregoing procedure will occurto those skilled in the art, and to the extent that such changes andmodifications are embraced by the appended claims, it is to beunderstood that they constitute part of my invention.

I claim:

1. The process which comprises reacting a tertiary amine salt ofpenicillin with an aqueous solution of an alkali forming hydroxideselected from the group consisting of alkali metal hydroxides andalkaline earth metal hydroxides in the presence of a substantiallywater-immiscible organic solvent and recovering the corresponding metalsalt of penicillin from the reaction mixture.

2. The process which comprises reacting a tertiary amine salt ofpenicillin with an aqueous solution of sodium hydroxide in the presenceof a substantially water-immiscible organic solvent and recovering thesodium salt of penicillin from the reaction mixture.

3. The process which comprises reacting the N-ethyl piperidine salt ofpenicillin with an aqueous solution of sodium hydroxide in the presenceof a substantially water-immiscible organic solvent and recovering thesodium salt of penicillin from the reaction mixture.

4. The process which comprises reacting the N-ethyl piperidine salt ofpenicillin with an aqueous solution of calcium hydroxide in the presenceof a substantially water-immiscible organic solvent and recovering thecalcium salt of penicillin from the reaction mixture.

. MAX TISHLER.

No references cited.

